Minitube of America will use SBIR funds to develop porcine models of atherosclerosis. Cardiovascular disease is the leading cause of morbidity and mortality in the United States and coronary artery disease accounts for a majority of the deaths. Research to uncover the etiology of atherosclerosis and to develop new therapies has relied heavily on the use of animal models. Most of these studies point to hypercholesterolemia as the principal cause of atherosclerosis. Rabbits, swine and rhesus monkeys with genetic mutations linked to hypercholesterolemia have been used to study atherosclerosis but recent research has focused on genetically modified mice. However, genetically modified mice that manifest hypercholesterolemia do not exhibit lesions typical of atherosclerosis in humans. In contrast, swine pedigrees that develop hypercholesterolemia as a result of naturally occurring genetic mutations manifest arterial lesions similar to those seen in humans. The goal of this project is to introduce specific genetic alterations in miniature swine that reproduce the genetic and pathological characteristics of human atherosclerosis. Studies in humans, mice and swine indicate that alterations in the low-density lipoprotein gene (Ldlr) can produce hypercholesterolemia and early onset atherosclerotic lesions in coronary and other arteries. The approach in this project will be to use traditional and/or novel gene targeting methods to knockout both alleles of Ldlr in male and female miniature swine cells. These Ldlr-/-cells will be used in nuclear transfer to produce Ldlr-/-founder animals that will be analyzed for their propensity to develop atherosclerosis. These genetically modified miniature swine will be expanded through breeding and then sold to meet a growing need of medical device and pharmaceutical companies for uniform animal models of human pathologies that can help predict the outcome of human therapeutic interventions. [unreadable] [unreadable]